The invention relates to a method for detecting or demonstrating the presence of molecules, in particular biomolecules, which are present in a liquid phase.
The detection of molecules, especially biomolecules contained in liquids, is becoming more and more important economically, especially in the fields of biology, chemistry, biochemistry, pharmaceutics, as well as medical diagnostic and research.
The following features are critical for the practical application of a method according to the invention:                a fast and specific detection;        a label-free detection;        the option of following time-dependent functions in real-time, e.g. for examining the kinetics of (bio) chemical processes;        the suitability for using it under reactive conditions;        the bio-compatibility; and        the omitting of toxic components.        
The following features are critical for using a device according to the invention in practical operations:                the easy handling;        the cost-effective implementation;        a compact design, preferably offering the option of miniaturization up to the micrometer range, meaning along length scales in the range of or smaller than the dimensions of biological cells;        the option of a detection inside closed (bio) chemical (reaction) vessels;        the ability to integrate into arrays consisting of many different sensors which preferably react specifically to different substances but can be read out with the same readout mimic;        the option of a fast, sequential and/or parallel readout of many sensors, especially via multi-channel detectors or with the aid of multiplexing;        the option for an in-situ examination.        
From a physical perspective, label-free biosensors are categorized into three groups according to the publication by Ivnitski, D; Abdel-Hamid, I.; Atanasov, P. and Wilkins, E., “Biosensors for detection of pathogenic bacteria,” published in Biosens. Bioelectron. 14, pp 599, 1999:                1. Mass-sensitive sensors with piezoelectric materials or cantilevers for detecting the presence of biological material;        2. Electro-chemical sensors which are based on measuring the current or the resistance of a transducer; and        3. Optical sensors which substantiate the change in the refractive index or of the presence of a thin layer.        
A biosensor is known from the U.S. Pat. No. 7,118,710 B2 which comprises a two-dimensional grating or grating with high refractive index, deposited on a substrate, as well as one or several specific, binding, label-free substances which are immobilized on the surface of the grating. A change in the diffraction intensity appears when illuminating the biosensor.
The International patent application Publication No. WO 2000/041213 A1 describes the dip-pen nanolithography (DPN) technique for producing structures having dimensions of 10-1000 nm by using an atomic force microscope for which the tip is wetted with a solution (ink) that is transferred by a driving force to the surface of a substrate. Owing to the humidity in the air, a watery meniscus forms between the tip and the substrate surface which functions to transfer the molecules from the tip to the surface where the molecules are absorbed chemically or where they arrange themselves on correspondingly prepared surfaces.
The European patent document EP 1881368 A2 discloses the use of membrane lipids as ink for use with the DPN, wherein the number of lipid (double) layers on the substrate surface can be adjusted via the speed of the tip on the atomic force microscope above the substrate surface and the relative humidity in the atmosphere.
The use of DPN for producing micro structures and nano structures from liquid membrane lipids which contain different shares of macro molecules and their characterization through fluorescence and photo emissions electron microscopy are described in Multiplexed Lipid Dip-Pen Nanolithography on Subcellular Scales for the Templating of Functional Proteins and Cell Culture, published by S. Sekula, J. Fuchs, S. Weg-Remers, P. Nagel, S. Schuppler, J. Fragala, N. Theilacker, M. Franzreb, C. Wingren, P. Ellmark, C. A. K. Borrebaeck, C. A. Mirkin, H. Fuchs and S. Lenhert, Small 4, pp 1785-1793, 2008.
In the publication Science 263, p. 60, 1994, A. Kumar and G. Whitesides describe the condensation of water on a self-organizing monolayer of ω functionalized alkane thiolates on gold. This condensation diagram forms an optical refractive grating for reflected or transmitted light with a wavelength of 632.8 nm which is subjected to an atmosphere with a constant relative humidity of the air.